Chlamydia trachomatis infections cause sexually transmitted disease (STD) and blinding trachoma; diseases that affect hundreds of millions of people world-wide. A pathognomonic feature of chlamydial infection is the failure to evoke adequate protective immunity resulting in re-infection or persistent infection. The overall goal of our work is to understand the pathogenic mechanisms by which chlamydiae evade host defenses and establish persistent infection. To this end, we have shown that the chlamydial plasmid is major virulence factor. Plasmid-free chlamydia produce attenuated self-limiting infections that generate minimal to no pathology. Notably, attenuated infection generates a robust B and T cell immunity that is protective following challenge with virulent plasmid positive organisms. To understand how the plasmid functions as a virulence factor we made deletion mutants for all 8-plasmid gene encoded proteins (pgp) and characterized the strains in in vitro and in vivo infection models. We found that pgp 1,2,5,6,7 and 8 have housekeeping functions that control plasmid replication and maintenance. In contrast, Pgp 3 and 4 were shown to be dominant virulence factors. We showed that pgp 4 is a transcriptional regulator of multiple chromosomal virulence genes that function in chlamydial exit from host cells and the establishment of persistent infection. These finding provide an explanation at the molecular level for the dominant infection attenuating phenotype of plasmid-free organisms. Thus, plasmid-free chlamydia are being investigated as live-attenuated vaccines for the prevention of STD and blinding trachoma. We are also pursuing live attenuated chlamydia carrying an engineered chlamydial plasmid with deleted virulence genes pgp3 and pgp4 capable of expressing non-chlamydial genes as a novel vaccine platform for the prevention of other important mucosal diseases.